I am running an analysis in CuffDiff using a GFF file of annotations from FlyBase (www.flybase.org). The GFF file contains annotation data for genes, mRNA, CDS and 5'- and 3'-UTR's.
Differential expression of isoforms is reported in some 1266 pairwise comparisons. However, most of the genes in the analysis contain only 1 transcript; and there is very little overlap between the genes showing differential expression (file gene_exp.diff) and the isoforms showing differential expression (file isoform_exp.diff). Can any one explain to me how this is possible?
My code is below:
cuffdiff \
-o "/gpfs/group1/f/flyinv/Outputs_CuffDiff/Exons_B_allFemales_Ch3/" \
-L AR,CH,PP,PX,ST,TL \
--total-hits-norm \
--dispersion-method per-condition \
--frag-bias-correct /gpfs/group1/f/flyinv/working_index/Dpse3_0_1.fa \
--multi-read-correct \
--library-norm-method geometric \
/gpfs/group1/f/flyinv/working_index/Ch3_FB_genes_exons_mRNA_CDS_UTR_PalmieriCh3Genes.gff3 \
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/AR_DM1005_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/AR_KB945_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/AR_MSH126_Female/accepted_hits.bam \
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/CH_JR272_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/CH_KB888_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/CH_MSH202_Female/accepted_hits.bam \
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/PP_DM1049_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/PP_DM1054_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/PP_DM1065_Female/accepted_hits.bam \
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/PX_SPE123_1-2_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/PX_SPE123_4-1_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/PX_SPE123_5-2_Female/accepted_hits.bam \
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/ST_JR138_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/ST_MSH177_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/ST_MSH49_Female/accepted_hits.bam \
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/TL_SCI_12-2_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/TL_SPE123_7-1_Female/accepted_hits.bam,\
/gpfs/group1/f/flyinv/Outputs_TopHat/transcriptomeSequence_exons/TL_SPE123_8-1_Female/accepted_hits.bam
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